Colonization of barley roots by Fusarium culmorum and influence of Pseudomonas fluorescens on the process

The stages of barley root colonization by Fusarium culmorum were studied in sterile vermiculite by the method of fluorescent antibodies. The influence of the antagonistic bacterium Pseudomonas fluorescens on the process of root colonization by F. culmorum was demonstrated. In vermiculite inoculated with F. culmorum, the fungus density on the roots increased gradually. In the case of joint inoculation of vermiculite with the fungus and the bacterium, the F. culmorum density on the roots changed abruptly. It was shown that the site of primary colonization of the roots by the fungus was mainly the zone of root hairs. When Pseudomonas fluorescens was present on the roots, F. culmorum colonized not only root hairs, but also the elongation zone, during the first two days. Introduction of Pseudomonas fluorescens into vermiculite resulted in lower intensity of barley root rot.     

Effects of Fusarium culmorum head blight on mycotoxin accumulation and yield traits in barley doubled haploids

The susceptibility of barley doubled haploids (DH) to Fusarium head blight (FHB) was investigated. Heads of 24 DH lines (11 two-rowed and 13 six-rowed) derived from F₁ Maresi (two-rowed) × Pomo (six-rowed) hybrids were inoculated with a conidial suspension of the single isolate IPO348-01 of Fusarium culmorum. The experiment was carried out in three consecutive years (1996–98) in one location. The number of kernels per ear, 1000-kernel weight and kernel weight per ear were recorded in inoculated and control plots. In the infected kernels nivalenol (NIV) content and deoxynivalenol (DON) content were determined. The effects of genotype, year and genotype-year interaction on reduction of yield traits were significant. For mycotoxin content only genotype and year effects were found to be important. The average NIV concentration in kernels of inoculated lines ranged from 0.15 mg/kg in the two-rowed line MP7 to 6.36 mg/kg in the six-rowed line MP113. A low accumulation of DON was observed in the studied population (from 0.01 to 0.20 mg/kg). Generally, no significant differences in mycotoxin content were found between two-rowed and six-rowed genotypes. The line MP7 was found to be superior — with the lowest mycotoxin accumulation, and reduction in yield traits. Environmental conditions (years) affected DON and NIV level in kernels; however, the tendency to a lower or higher accumulation of mycotoxin in individual lines was stable over the years.     

Formation of moniliformin by Fusarium sporotrichioides and Fusarium culmorum

Two strains of Fusarium sporotrichioides and one strain of F. culmorum were shown to produce the mycotoxin moniliformin in rice culture. Identification was by reverse-phase liquid chromatography, thin-layer chromatography, and mass spectrometry.     

Contamination of malt barley and wheat by Fusarium graminearum and Fusarium culmorum from the crop years 2001-2003 in eastern Croatia

This study investigated infection levels with Fusarium graminearum and Fusarium culmorum in malt barley and wheat in eastern Croatia. The contamination was surveyed over three consecutive crop years (2001–2003) on five locations for barley and three wheat cultivating locations. F. graminearum loads reached levels of potentially serious threat for the commercial production of malting raw materials in both cereals (up to 29.1%). On the other hand, the mean percentage of kernels infected with F. culmorum was low to medium (up to 6.1%). The fungal invasions for years and locations were affected by meteorologic and other environmental factors and the pattern seemed to be consistent with species-specific optimal conditions reported by other authors.     

Formation of moniliformin by Fusarium sporotrichioides and Fusarium culmorum.

Two strains of Fusarium sporotrichioides and one strain of F. culmorum were shown to produce the mycotoxin moniliformin in rice culture. Identification was by reverse-phase liquid chromatography, thin-layer chromatography, and mass spectrometry.     

Effect of high-lysine mutations on the protein fractions of barley grain

We have studied the effects of six high-lysine barley mutations (Risø mutants 1508 and 56, Notch 1 and 2, lys 95 and 449) on the protein fractions of the grain. All mutants had a decreased relative and total amount of the lysine-poor hordein fraction, but only in Risø 1508 and 56 was the polypeptide composition of this fraction greatly affected. In all the mutants there were increases in the lysine-rich glutelin proteins and in nonprotein N while in Risø 1508 and the Notch mutants the total amount of salt-soluble proteins was also increased and their relative polypeptide composition substantially altered.This work was supported by EEC Contract No. 473 under the common program on plant protein improvement.     

Evaluation of Spring and Winter Wheat Reaction to Fusarium culmorum and Fusarium avenaceum

Winter (37), spring (8) wheat accessions and additionally, 7 double haploid (DH) lines were examined for susceptibility to Fusarium seedling blight after inoculation with F. culmorum and F. avenaceum. Winter accessions exhibited lower susceptibility of about 30% to both pathogens than spring cultivars. Susceptibility of winter cultivars varied from low (22%) to high (97%). Evaluation of the root was found to be more reliable than evaluation of coleoptile necrosis.
F. avenaceum infected mostly root and, to a lesser extent, coleoptile and leaves, with about a three times lower disease score of coleoptile against root. F. culmorum caused a 1.5 higher disease score on root than on coleoptile. Susceptibility of DH lines was different from susceptibility of parental forms. Reaction of individual accessions to F. culmorum and F. avenaceum was different.     

Production and secretion of 5-n-alkylresorcinols by Fusarium culmorum

Fusarium culmorum F1 was found to produce and secrete into the culture medium several of 5-n-alkylresorcinols. The amount of resorcinolic lipids was 5.3 microg/g and 0.9 microg/l in mycelium and in post-culture liquid, respectively. First of all F. culmorum F1 produces saturated homologues with C15 to C25 side chains. The extract from the medium contained only homologues with shorter carbon chains (C13 to C17).     

Comparison of Fusarium acuminatum and Fusarium culmorum isolates by means of tandem-crossed immunoelectrophoresis

F. acuminatum and F. culmorum strains were compared by means of tandemcrossed immunoelectrophoresis in order to estimate the possiblities of serological classification in Fusarium sections Gibbosum and Discolor. On the basis of qualitative similarity the two species could be distinguished well. By the use of anti-F. acuminatum serum a similarity of S_SM=0.52 was found between F. acuminatum and F. culmorum, but the S_SM coefficient reached a value of 0.67 when the anti-F. culmorum serum was tested. This asymmetric nature of the qualitative similarity is discussed. In the majority of cases the quantitative differences of the common antigens did not allow differentiation between the species.     

The effect of virus infection on morphology and protein components of pollen grains

Morphology of pollen grains collected from healthy and virus infected plants ofChenopodium quinoa L.,Chenopodium album L. andNicotiana tabacum L. cv. Samsun was investigated using scanning electron microscopy (SEM). Pollen grains from tobacco plans infected with tobacco mosaic virus (TMV) were smaller, with rounded shape and conspicuous deformation of aperture unlike oval and smooth pollen grains from healthy plants. No morphological alterations were observed inC. quinoa andC. album plants infected with TMV and cucumber mosaic virus (CMV). Polyacrylamide gel electrophoresis of pollen proteins revealed substantial quantitative and qualitative differences in protein components of pollen grains collected from healthy and virus infected plants     

PCR detection assays for the trichothecene-producing species Fusarium graminearum, Fusarium culmorum, Fusarium poae, Fusarium equiseti and Fusarium sporotrichioides

Contamination of small-grain cereals with the fungal species Fusarium graminearum, F. culmorum, F. poae, F. sporotrichioides and F. equiseti is an important source of trichothecenes, Zearalenone and other mycotoxins which cause serious diseases in human and animals. Additionally, these species contribute to Fusarium Head Blight, a disease which produces important losses in cereal yield. Early detection and control of these Fusarium species is crucial to prevent toxins entering the food chain and a useful tool in disease management practices. We describe the development of specific PCR assays to F. graminearum, F. culmorum, F. poae, F. sporotrichioides and F. equiseti using DNA from pure fungal cultures as well as from naturally infected wheat seeds, using in this case a rapid and easy protocol for DNA isolation. The specific primers were designed on the basis of IGS sequences (Intergenic Spacer of rDNA), a multicopy region in the genome that permits to enhance the sensitivity of the assay in comparison with PCR assays based on single-copy sequences.     

Analysis of expressed sequence tags from cDNA library of Fusarium culmorum infected barley (Hordeum vulgare L.) roots

Fusarium culmorum is one of the most common and globally important causal agent of root and crown rot diseases of cereals. These diseases cause grain yield loss and reduced grain quality in barley. In this study, we have analyzed an expressed sequence tag (EST) database derived from F. culmorum infected barley root tissues available at the National Center for Biotechnology Information (NCBI). The 2294 sequences were assembled into 1619 non-redundant sequences consisting of 359 contigs and 1260 singletons using the program CAP3. BLASTX analysis for these sequences was conducted in order to find similar sequences in all databases. Gene Ontology search, enzyme search, KEGG mapping and InterProScan search were done using Blast2GO 3.0.7 tool. By BLASTX analysis, 41.7%, 7.7%, 3.2% and 47.4% of ESTs were categorized as annotated, unannotated, not mapping and without blast hits, respectively. BLASTX analysis revealed that the majority of top hits were barley proteins (43.5%). Based on Gene Ontology classification, 38.3%, 31.3%, and 16% of ESTs were assigned to molecular function, biological process, and cellular component GO terms, respectively. Most abundant GO terms were as follows: 157 sequences were related to response to stress (biological process), 207 sequences were related to ion binding (molecular function), and 160 sequences were related to plastid (cellular component). Furthermore, based on KEGG mapping, 369 sequences could be assigned to 264 enzymes and 83 different KEGG pathways. According to Enzyme Commission (EC) distribution; 94 sequences were transferases (EC2) while 70 sequences were hydrolases (EC3).     

Arabidopsis is susceptible to the cereal ear blight fungal pathogens Fusarium graminearum and Fusarium culmorum

The fungal pathogens Fusarium graminearum and F. culmorum cause ear blight disease on cereal crops worldwide. The disease lowers both grain quality and grain safety. Disease prevalence is increasing due to changes in cropping practices and the difficulties encountered by plant breeders when trying to introgress the polygene-based resistance. The molecular basis of resistance to Fusarium ear blight in cereal species is poorly understood. This is primarily due to the large size of cereal genomes and the expensive resources required to undertake gene function studies in cereals. We therefore explored the possibility of developing various model floral infection systems that would be more amenable to experimental manipulation and high-throughput gene function studies. The floral tissues of tobacco, tomato, soybean and Arabidopsis were inoculated with Fusarium conidia and this resulted in disease symptoms on anthers, anther filaments and petals in each plant species. However, only in Arabidopsis did this initial infection then spread into the developing siliques and seeds. A survey of 236 Arabidopsis ecotypes failed to identify a single genotype that was extremely resistant or susceptible to Fusarium floral infections. Three Arabidopsis floral mutants that failed to develop anthers and/or functional pollen (i.e. agamous-1, apetala1-3 and dad1) were significantly less susceptible to Fusarium floral infection than wild type. Deoxynivalenol (DON) mycotoxin production was also detected in Fusarium-infected flowers at >1 ppm. This novel floral pathosystem for Arabidopsis appears to be highly representative of a serious cereal crop disease.     

The influence of gamma radiation and substrate on mycotoxin production by Fusarium culmorum IMI 309344

Mycotoxin production (deoxynivalenol (DON), acetyl deoxynivalenol (A DON) and zearalenone) by Fusarium culmorum inoculated on to maize (heat sterilized, irradiation sterilized and non-sterile) and irradiated to 1 kGy or 3 kGy, or unirradiated, was investigated over a period of time. Lowest mycotoxin production was observed on non-sterile maize which may be due to the presence of a competitive microflora on non-sterile maize. In general, mycotoxin production was higher on heat-sterilized grain as compared to irradiation-sterilized maize. It was suggested that this pattern of mycotoxin production was possibly caused by changes in the grain brought about by autoclaving, which favoured mycotoxin production and possibly induced changes in irradiation-sterilized maize which inhibited mycotoxin production. On sterile maize, there was no significant difference in DON production by unirradiated, 1 kGy and 3 kGy irradiated cultures up to 56 d of incubation; between days 56 and 77 of incubation, DON production increased rapidly with largest increases occurring in irradiated (1 kGy and 3 kGy) cultures. On non-sterile grain, neither DON nor A DON were detected in unirradiated cultures of F. culmorum but were detected in cultures irradiated to 1 kGy and 3 kGy. In practice grain should be stored under conditions of temperature and moisture content which prevent fungal growth. However, in this study, the grain was stored under conditions that were approaching ideal for growth of the test organism. The results highlight that irradiation disinfestation of grain must be combined with good grain handling practices so that excessive mycotoxin production can be prevented during storage.     

Effect of phytotoxic metabolites ofFusarium culmorum on barley root and root-hair development

Summary A mixture of gluconic acid and two unidentified organic acids, isolated from the culture filtrate ofFusarium culmorum, grown in glucose-limited continuous culture, were found to be highly toxic to growth of barley seedlings. A concentration of 900 g ml^–1 of the acid totally inhibited germination of barley seeds, with lower concentrations greatly inhibiting growth and development of germinated seeds. Root extension and root-hair production were inhibited at a concentration of 450 g ml^–1 of the acids, but the number of roots per seedling was increased. Toxicity of the acids was promoted at low pH.     

Production of trichothecene mycotoxins byFusarium graminearum andFusarium culmorum on barley and wheat

Wheat cultivars (Stoa, MN87150, SuMai-3, YMI-6, Wheaton) and barley cultivars (Robust, Excel, Chevron, M69) were inoculated in the field with isolates ofFusarium graminearum andF. culmorum. The diseased (Fusarium head blight) kernels were analyzed for deoxynivalenol (DON), 15-acetyldeoxynivalenol (15-ADON) and nivalenol (NIV).F. culmorum produced all three trichothecenes on all cultivars tested whereasF. graminearum only produced DON and 15-ADON. There was no well defined correlation between DON production in the host and resistance although the data tended to favor SuMai-3 as having definitive resistance to bothF. graminearum andF. culmorum.Minnesota Agricultural Experiment Station, Paper No. 20 279.